Medical Sciences Commons^™

Tetrahydroisoquinoline Neurotoxins In Parkinson Disease, Michael G. Decuypere

Theses and Dissertations (ETD)

The goal of this dissertation work was to (1) determine the distribution of several tetrahydroisoquinoline (TIQ) derivatives in rodent, normal human and Parkinson disease (PD) brain, (2) quantify the levels of these TIQ derivatives in common food sources in an effort to link specific food intake patterns with the development of PD and (3) examine the neurotoxicity of select TIQ derivatives in human dopaminergic cell culture. The TIQs are a family of monoamine alkaloids that share structural homology with 1-methyl-4-phenyl-1,2,3,6-tetrahyrdropyridine (MPTP), can be formed from dopamine or its oxidized metabolites and may be involved in the pathogenesis of monoaminergic cell …

H5n1 Influenza Virus Induces A Parkinsonian Pathology, Haeman Jang

Theses and Dissertations (ETD)

The greatest threat for an influenza pandemic at this time is posed by the highly pathogenic H5N1 avian influenza virus. To date, 63% of the 436 known human cases of H5N1 infection have proven fatal. Animals infected by H5N1 viruses have demonstrated acute neurological signs ranging from mild encephalitis to motor disturbances and coma. However, no studies have examined the longer-term neurologic consequences of H5N1 infection. We show that this virus travels from the peripheral nervous system into the central nervous system (CNS) to higher levels of the neuroaxis, using C57BL/6J mice that are infected by the A/VN/1203/04 H5N1 virus …

T-Cell Receptor Cdr3 Sequence But Not Recognition Characteristics Distinguish Autoreactive Effector And Foxp3+ Regulatory T-Cells, Xin Liu

Theses and Dissertations (ETD)

Regulatory T-cells (Treg) play an important role in maintaining immune tolerance to self-antigens and in suppressing excessive immune responses in the host. The source, specificity, and plasticity of the forkhead box transcription factor 3 (Foxp3⁺) Treg and conventional T (Tconv) cell populations active at sites of autoimmune pathology are not well characterized. To evaluate this, we combined global repertoire analyses and functional assessments of isolated T-cell receptors (TCR) from TCRa retrogenic mice with autoimmune encephalomyelitis (EAE). Treg and Tconv cell TCR repertoires were distinct, and autoantigen-specific Treg and Tconv cells were enriched in diseased tissue. Autoantigen sensitivity and …

Glycan Shielding Of The Influenza Virus Hemagglutinin Elicits Evasion Of The Adaptive Immune Response And T-Cell-Driven Pathology, Keith C. Wanzeck

Theses and Dissertations (ETD)

Three separate influenza pandemics have emerged in the human population since 1918, each characterized by viruses that lack N-linked glycosylation sites on the globular head of the hemagglutinin protein. In contrast, recent non-pandemic isolates have acquired such sites. Here we constructed isogenic viruses containing differing numbers of additional N-linked glycosylation sites to assess the impact on the host immune response. These studies show that mice infected with a glycosylated virus remain susceptible to challenge with a non-glycosylated virus, glycosylated viruses elicit an inferior immune response, and in this context T-cell pathology and death may occur. We conclude from these data …

Ns1 Of H5n1 Interacts With Sap-97 In A Pdz-Dependent Manner To Disrupt Epithelial Barrier Integrity, Veronica Garcia Conoley

Theses and Dissertations (ETD)

The ability of influenza A virus to cause global pandemics has been a great concern throughout history and poses a serious health risk worldwide. Pandemic outbreaks throughout history, such as the Spanish flu of 1918, have claimed the lives of millions of people worldwide. The current outbreak of avian influenza (H5N1) that began in 1997 is still claiming lives, and therefore efforts to understand the mechanisms of pathogenesis in this highly virulent virus are of the utmost importance. According to the World Health Organization, there have been 447 reported H5N1 human cases, resulting in 263 deaths. The pathology of H5N1 …

Structural Characterization Of The C-Terminus Of The Cochlear Motor Protein Prestin: Intrinsic Disorder Linked To Voltage Sensor Function, Michael S. Podgorski

Theses and Dissertations (ETD)

The sensitivity of mammalian hearing relies upon the ability to amplify sound. Outer hair cells (OHCs) in the cochlea have motile properties that support this function. The motor protein called prestin resides in the lateral wall of the OHCs. Prestin undergoes voltage-dependent conformational changes, which correlates to the movement of charge through the membrane or non-linear capacitance (NLC). This property underlies OHC electromotility. A mechanistic understanding of prestin’s function remains unknown. Previous research has tied prestin’s C-terminus (residues 499-799) to the voltage-dependent events that occur in the membrane. The folding and biochemical properties of prestin’s C-terminus were characterized. This information …

Discovery And Validation Of New Regulatory Rna Elements In Chlamydia Trachomatis, Yasser Mohammed Elsayed Metwally Abdelrahman

Theses and Dissertations (ETD)

Chlamydia trachomatis is an obligate intracellular bacterium that exhibits a unique biphasic developmental cycle that can be disrupted by growth in the presence of IFN-g and b-lactams, giving rise to an abnormal growth state termed persistence. Relatively little is known about the regulatory mechanisms that control temporal gene expression during the developmental cycle or the control of persistence and reactivation. Here we have examined the expression of a newly defined family of non-coding RNAs (ncRNAs) that are differentially expressed during the developmental cycle and the induction of persistence and reactivation (Using IFNγ and Carbenicillin). Non-coding RNAs were initially identified using …

Identification Of The Downstream Effector Genes Involved In Hoxb4-Induced Expansion Of Hematopoietic Progenitor Cells, Jie Jiang

Theses and Dissertations (ETD)

Overexpression of HOXB4, a member of Homeobox transcription factor family, promotes expansion of hematopoietic stem and progenitor cells bothin vivo and in vitro. However, the molecular mechanisms underlying this effect are not well understood. In order to identify direct target genes of HOXB4 in primary murine hematopoietice progenitor cells, we induced HOXB4 function in lineage-negative, murine bone marrow cells, using a tamoxifen-inducibleHOXB4-ER^T2fusion protein. Seventy seven genes with differentially changed expression in early response to HOXB4 have been identified as candidate target genes. Among them, we show that Hemogen (Hemgn), encoding a nuclear protein specifically …

Pandemic Potential Of Reassortant Swine Influenza A Viruses, Christy Brockwell Staats

Theses and Dissertations (ETD)

Influenza A viruses are capable of causing disease in several species, including birds, humans and swine. Host specificity of the viruses is not absolute, and is influenced by a range of factors. Swine play a pivotal role in the interspecies transmission of influenza A viruses, as they are susceptible to infection with both human and avian strains and have been implicated as a “mixing vessel” for the reassortment of influenza A viruses from different species. The reassortment of influenza A viruses of human and avian origin led to human influenza pandemics in 1957 and 1968.

The dynamics of swine influenza …

Distinct Dna Damage Signaling In The Brain Distinguishes Atld, Nbs, And Atr-Seckel Syndrome, Erin Renee Phillips Shull

Theses and Dissertations (ETD)

DNA double strand breaks create a situation of extreme stress under which a cell must either be capable of repairing the lesions in order to continue replication or succumb to death. Not surprisingly, deficiencies in DNA repair genes often lead to human diseases frequently associated with genomic instability, cancer proneness, and neuropathology. Neurological consequences of aberrant DNA repair mechanisms vary depending upon the affected gene and the pathway in which it operates. Ataxia-telangiectasia (A-T) is the prototypical disease associated with DNA double strand break (DSB) repair deficiency and is characterized by severe neural pathology. A-T results from homozygous mutations that …

Regulation Of Pyruvate Dehydrogenase Kinase 4 By Thyroid Hormone / Role Of Peroxisome Proliferator Activated Receptor Gamma Coactivator-1 Alpha And Ccaat Enhancer Binding Protein, Ramy Naguib Attia

Theses and Dissertations (ETD)

Pyruvate dehydrogenase kinase 4 (PDK4) regulates pyruvate oxidation through the phosphorylation and inhibition of the pyruvate dehydrogenase complex (PDC). The PDC catalyzes the conversion of pyruvate to acetyl-CoA and it is an important control point in glucose and pyruvate metabolism. Previous studies had reported that PDK4 gene expression is induced by thyroid hormone (T3). These studies did not investigate the mechanisms by which T3 regulated PDK4 gene expression. I have examined the role of the thyroid hormone receptor (TR), transcriptional coactivators especially the peroxisome proliferator activated receptor gamma coactivator-1 (PGC-1α) and other transcription factors that act as accessory factors in …

Spatial Resolution Analysis Of A Variable Resolution X-Ray Cone-Beam Computed Tomography System, Bahram Dahi

Theses and Dissertations (ETD)

A new cone-beam computed tomography (CBCT) system is designed and implemented that can adaptively provide high resolution CT images for objects of different sizes. The new system, called Variable Resolution X-ray Cone-beam CT (VRX-CBCT) uses a CsI-based amorphous silicon flat panel detector (FPD) that can tilt about its horizontal (u) axis and vertical (v) axis independently. The detector angulation improves the spatial resolution of the CT images by changing the effective size of each detector cell. Two components of spatial resolution of the system, namely the transverse and axial modulation transfer functions (MTF), are analyzed in …

Contribution Of Organic Cation Transporter 2 (Oct2) To Cisplatin-Induced Nephrotoxicity, Kelly K. Filipski

Theses and Dissertations (ETD)

Cisplatin is the most widely used anticancer agent; however, the cellular pharmacokinetics are poorly understood. Cisplatin is predominantly eliminated through the urine via active secretion and is associated with nephrotoxicity. Currently, prehydration therapy is employed to prevent toxic renal side effects; however it has not been completely ameliorated. The studies described herein aim to determine the mechanism in which cisplatin enters the kidney cell from the blood and how it is subsequently secreted into the urine. Organic cation transporter 2 (OCT2) and ABCC2 are highly expressed in the kidney on the basolateral and apical membrane, respectively. We determined the contribution …

Generation And Characterization Of A Knock-In Allele Of Eklf: Probing The In Vivo Role Of The Chromatin Remodeling Domain In Definitive Hematopoietic Cells, Valerie Malyvanh Jansen

Theses and Dissertations (ETD)

The zinc finger-encoding transacting factor EKLF, or erythroid Krüppel-like factor, binds key regulatory elements of many erythroid-specific genes, and is essential for definitive erythropoiesis. Mice lacking this factor die of anemia by E15.5 of gestation, failing to activate β-globin gene transcription, and demonstrating a block in the erythroid differentiation program at the primitive erythroblast stage. In contrast, megakaryocytic progenitors are amplified in EKLF-null embryos, with increased Fli-1 gene expression, a marker of early megakaryocytic differentiation. These observations are consistent with the idea that EKLF modulates the megakaryocytic-erythroid (M-E) differentiation switch.

Our laboratory has previously demonstrated that an amino terminal sequence …

Studying The Specificity Of Hpxr Antagonists Using A Panel Of Cell-Based Assays, Fang Lei

Theses and Dissertations (ETD)

The pregnane X receptor (PXR or SXR; NR1I2) is a member of the nuclear receptor superfamily. It activates the transcription of a large network of genes including cytochrome P450 (CYP) and Mdr1 which play critical roles in chemicals metabolism and transportation. Induction of CYPs contributes to adverse drug-drug interactions. Non-toxic, PXR-specific antagonists will be valuable in attenuating the adverse drug-drug interaction which is the cause of many treatment failures in clinic. However, few hPXR antagonists were reported particularly the specific ones. In this thesis a reporter gene assay was used to study the specificity of hPXR antagonists from a panel …

Multiscale Genomic Analysis Of The Corticolimbic System: Uncovering The Molecular And Anatomic Substrates Of Anxiety-Related Behavior, Khyobeni Mozhui

Theses and Dissertations (ETD)

Genetic diversity generates variation at multiple phenotypic levels, ranging from the most basic molecular to higher-order cognitive and behavioral traits. The far-reaching impact that genes have on higher traits is apparent in several neuropsychiatric conditions such as stress and anxiety disorders. Like most, if not all, neural phenotypes, stress, anxiety, and other emotion-related traits are extremely complex and are defined by the interplay of multiple genetic, environmental, experiential, and epigenetic factors.

The work presented in this dissertation is a multi-scalar, integrative analysis of the molecular and neuroanatomic substrates that underlie emotion-related behavior. The amygdala is a principle component of the …

Pax5 Haploinsufficiency Cooperates With Bcr-Abl1 To Induce Acute Lymphoblastic Leukemia, Christopher B. Miller

Theses and Dissertations (ETD)

Acute lymphoblastic leukemia (ALL) is the commonest pediatric malignancy and comprises several distinct subtypes each with its own unique pathogenesis, clinical behavior, and response to therapy. Chromosomal aberrations are a hallmark of ALL but alone fail to induce leukemia. Pediatric ALLs can be divided into several categories based on the expression of several genetically conserved chromosomal translocations including the t(9,22)[BCR-ABL1], t(1,19)[TCF3-PBX1], t(12,21)[ETV6-RUNX1], MLLrearranged leukemia’s, hyperdiploid and hypodiploid karyotypes, and T-lineage leukemia. Each translocation confers a characteristic transforming phenotype within the cell in which it originates but is alone insufficient to induce overt leukemia. …

The Combined Effect Of In-Situ Tumor And Irradiation On Peritumoral Brain Vasculature, Janice Ann Zawaski

Theses and Dissertations (ETD)

In the USA, 200,000 brain tumors are diagnosed each year with glioma representing 8.4% of the 200,000. The standard treatment for glioma consists of surgical resection, when possible, followed by radiation therapy (RT) and/or chemotherapy. Radiation therapy is one of the most effective treatments of brain tumors; however, the therapeutic ratio of RT is limited by damage to the normal tissue. We hypothesize that tumor growth has an adverse effect on the peritumoral tissue through the angiogenic/inflammatory environment it creates rendering it susceptible to further damage by RT which may be prevented by using anti-angiogenic/anti-inflammatory agents. We have developed a …

Insights Into Btb-Cul3 Ubiquitin Ligases From The Structures Of Spop-Substrate Complexes, Min Zhuang

Theses and Dissertations (ETD)

Cullin-Ring ubiquitin ligases (CRLs) are E3 complexes that specifically recognize substrates through substrate adaptors. In the largest CRL subfamily, Cul3 binds a BTB domain, and a protein-interaction domain such as MATH recruits substrates for ubiquitination. Here we present biochemical and structural analyses of the MATH and BTB domain containing protein, SPOP, which regulates diverse signaling pathways. First, we identified a conserved SPOP Binding Consensus (SBC) motif in the transcriptional regulator Ci, the protein phosphatase Puc, and the chromatin component MacroH2A. The SBC motif specifically binds the MATH domain of SPOP, and is required for Puc ubiquitination in vitro and in …

Structural And Kinetics Studies Of The Enzyme Dihydropteroate Synthase And The Implications For Antibiotic Resistance, Katherine A. Ayers

Theses and Dissertations (ETD)

The goal of this study is to develop small molecule inhibitors of DHPS for the treatment of a variety of infectious diseases. According to the World Health Organization (WHO), infectious diseases kill more than 13 million people worldwide every year making it the second leading cause of death behind cardiovascular disease. The sulfonamide class of drugs has been in use since the 1930’s to treat many infectious agents and act by targeting the enzyme dihydropteroate synthase (DHPS) of the prokaryotic and lower eukaryotic folate pathway.

DHPS is an ideal drug target because humans do not synthesize folate de novo and …

Abcb6 Is A Porphyrin Transporter With A Novel Trafficking Signal That Is Conserved In Other Abc Transporters, Yu Fukuda

Theses and Dissertations (ETD)

The ATP-binding cassette (ABC) transporters play an important role as a barrier to protect cells from the accumulation of toxic xenobiotics and metabolites due to their ability to translocate a wide array of compounds across lipid bilayers. However, many ABC transporters, especially the ones localized in the intracellular organelles, are involved in critical biological processes such as antigen presentation. The core unit of ABC transporters contains two functional domains: the membrane spanning domain (MSD) and the nucleotide binding domain. The full transporters contain two of these units in tandem in a single polypeptide, whereas the half transporters only contain one …

Torsina And The Pathophysiology Of Dyt1 Dystonia, Yu Zhao

Theses and Dissertations (ETD)

The goal of my dissertation work was to examine the systems biology of torsinA, a DYT1 dystonia-associated protein, by using rodent model systems. TorsinA is a putative ATPase associated with a variety of cellular activities (AAA+). Deletion of glutamic acid residue 302/303 in TOR1A is causally associated with many cases of early-onset primary dystonia.

In our work, transient forebrain ischemia and sciatic nerve transection were used as central and peripheral neural perturbations, respectively, to gain insight into the in vivo role(s) of torsinA. Moreover, transgenic mouse models that overexpress either human mutant torsinA (hMT) or wild-type torsinA (hWT) were used …

Regulation Of The Human Parainfluenza Virus (Hpiv3) Fusion Protein, Amanda Ruth Chapman

Theses and Dissertations (ETD)

Paramyxoviruses include a number of important human pathogens, including measles virus, mumps virus, and the human parainfluenza viruses (hPIV) 1-4, as well as several animal pathogens, such as Sendai virus, Nipah virus and Hendra virus. The creation of effective drugs and vaccines against this family of viruses would play an important role in decreasing the prevalence of these viruses and contributing to the health of both humans and animals worldwide. The purpose of this work was to determine how the fusion (F) protein is regulated with a focus on the heptad repeat B (HRB) region of the F protein located …

The Role Of Fgf Signaling In Retinal Development, Abbie A. Hartge

Theses and Dissertations (ETD)

The development of the retina is a precise balance between intrinsic competence and extrinsic factors. This interplay is known to regulate the generation of cell types in the developing retina and similar mechanisms have been found in other regions of the CNS. In the developing retina, FGFs are a large family of secreted polypeptide growth factors. Fgf15 is the major Fgf expressed during retinal development in mice. Fgf15 is an example of an FGF that has been shown to control proliferation, cell fate specification, differentiation and migration during development. In this thesis I used analysis of specific genes throughout retinal …

Role Of The Lpa2 Receptor In Protecting Against Apoptosis, Shuyu E

Theses and Dissertations (ETD)

Lysophosphatidic acid (LPA) is a naturally occurring lipid mediator. It exists abundantly in biological fluids such as serum, saliva, follicular fluid, seminal fluid and malignant effusions and induces a vast array of biological responses affecting cell growth, survival, differentiation, migration and morphology. We recently identified lysophosphatidic acid (LPA) as a potent antiapoptotic agent for the intestinal epithelium. Based on computational modeling octadecenyl thiophosphate (OTP) was synthesized: a novel rationally designed, metabolically stabilized LPA mimic. OTP was more efficacious than LPA in reducing g-irradiation-, camptothecin-, or TNF-a/cycloheximide-induced apoptosis and caspase 3, 8 and 9 activity in the IEC-6 cell line. The …

Studies To Characterize The Requirements For The Binding And Release Of Erdj3, A Mammalian Er Dnaj Homolog, From Substrates, Yi Jin

Theses and Dissertations (ETD)

Heat shock protein 70s (Hsp70s) and their DnaJ co-factors exist in all organisms and in all eukaryotic organelles. These highly conserved chaperone pairs facilitate a large number of cellular processes. ERdj3 was identified as a soluble, lumenal DnaJ family member that binds to unassembled immunoglobulin heavy chains (HC) along with the BiP chaperone complex in the endoplasmic reticulum of mammalian cells. Here we demonstrate that ERdj3 binds directly to two unfolded substrates: immunoglobulin γHC and denatured firefly luciferase. Using mutagenesis studies on ERdj3 in both in vivo and in vitro binding assays, we defined ERdj3’s critical amino acids in domain …

The Tetraspanin Cd9 Localizes To Platelet-Platelet Contacts And Regulates Thrombus Stability, Sarah Kathleen Hill

Theses and Dissertations (ETD)

CD9, a member of the tetraspanin family, is highly expressed on platelets (50,000-80,000 copies per platelet). Tetraspanins have been implicated in modulation of integrin function, and it is hypothesized that CD9 will modulate GPIIb-IIIa, the major platelet integrin.

The association between CD9 and GPIIb-IIIa was analyzed using immunoprecipitations and confocal microscopy. These two proteins were found to associate with each other, particularly at areas of platelet-platelet contact and at the periphery. Confocal analysis revealed CD9 localization was most intense at platelet-platelet contact, as well as in platelet filopodia and lamellipodia, but there is a lack of CD9 at areas of …

Interaction Of Bacteriophage Mu Middle Transcription Activator Protein Mor With Promoter Dna, Kartik Iyer

Theses and Dissertations (ETD)

Gene expression during lytic development of bacteriophage Mu is regulated by a

transcriptional cascade in three phases: early, middle and late. Transcription from the middle promoter Pm requires the 129-amino acid transcriptional activator Mor, a product of early transcription, and the Escherichia coli RNA polymerase. The Pm promoter has a recognizable -10 hexamer but lacks a -35 hexamer. Mor binds as a dimer to an imperfect dyad-symmetrical element containing two 6-bp inverted repeats and centered at -43.5 in Pm. The goals of this study were:

1. To test the prediction from the crystal structure of Mor that residues Y70 and …

Factors Influencing Topotecan Cns Penetration In Mouse Models, Jun Shen

Theses and Dissertations (ETD)

Camptothecin analogs such as topotecan are currently tested in clinical trials for brain tumors. However the clinical outcome is far below the expectations, which are derived from the promising preclinical studies. This discrepancy could be partially attributed to the presence of two barrier systems in the brain, the blood brain barrier (BBB) and blood cerebrospinal fluid (CSF) barrier (BCB). Further investigations have demonstrated multiple ATP-binding cassette (ABC) transporters are present at the two barriers. Camptothecin analogs are well known substrates for several of the transporters. This work defined the role of two ABC transporters, Bcrp1 and P-gp in determination of …

The Unfolded Protein Response Increases Production Of Pro-Angiogenic Factors By Tumor Cell Lines, Nan Liao

Theses and Dissertations (ETD)

The rapid growth and proliferation of tumor cells will be limited at a stage when they encounter inadequate levels of oxygen and nutrient supply within the poorly vascularized tumor mass. These severe conditions negatively affect the proper folding of nascent proteins in the endoplasmic reticulum (ER) and lead to accumulation of unfolded protein within ER which is referred to as ER stress. Consequently, it will trigger the unfolded protein response (UPR) signal pathway through ER membrane stress sensor proteins including activating transcription factor 6 (ATF6), inositol-requiring 1 (IRE1) and PKR-like ER localized kinase (PERK). The UPR is largely a cytoprotective …